Objectives: Multiple sclerosis (MS) is characterized by recurrent and progressive demyelination/remyelination cycles which result in development of scleroses (scars) in white and gray matter of CNS, axonal damage and neuronal loss. There is no cure for MS, and only symptomatic treatments are available.1 Chronic cuprizone exposure produces progressive demyelination in mice brain which reverses upon its removal from the diet.2 Since there is limited evidence of altered arachidonic acid (AA) metabolism in MS, we hypothesized that cuprizone exposure would alter the expression of genes involved in AA metabolism, and that these genes represent potential therapeutic targets. Methods: Adult C57B16 mice were exposed to cuprizone for 6 weeks and then returned to a normal diet. Demyelination was assessed by histochemistry and magnetic resonance imaging. Real-Time PCR and western blotting were used to determine gene and protein expression, respectively, of critical enzymes involved in AA metabolism. Results: We demonstrated that the Magnetization Prepared Rapid Gradient Echo (MPRAGE) is a useful MRI technique to assess quantitatively structural changes in myelin rate in the corpus callosum during cuprizone exposure. Histochemistry with the myelin stains Black Gold and Fluoromyelin demonstrated that frank demyelination and influx of glial cells into the corpus callosum begins at week 3 and peaks at week 5. However, a decrease in myelin and oligodendrocyte markers, myelin basic protein (MBP) and 2,3-cyclic nucleotide 3-phosphodiesterase (CNPase), was evident at week 1. Increased expression of CD11b and glial acidic fibrillary protein (GFAP), evidence of activated microglia and astrocytes, was also observed at week 1. Coincident with these early changes, we found an increase in cyclooxygenase (COX)-2 that persisted throughout the demyelination process as well as lipoxygenase 15-LO which was increased at week 1 and peaked in the height of frank demyelination suggesting that these AA metabolism genes are either involved in or respond to the earliest sign of demyelination. Moreover, western blotting analysis showed that 15-LO protein levels, peaked in the height of frank demyelination. Both gene and protein expression of 5-LO was not significantly changed during the early stages of demyelination but it peaked during week 5, when glial markers and frank demyelination also reached their peak of expression. While expression of 12-LO was not consistently increased during demyelination, increased 12-LO expression was observed during the remyelination, suggesting a role for this isoform in the recovery process. The expression cPLA2 and sPLA2 also increased at the peak of frank demyelination, while Ca2+-independent iPLA2 expression was not changed. Conclusions: Multiple enzymes involved in AA metabolism are altered in the cuprizone model of MS suggesting that COX- and LO- derived AA metabolites are involved in demyelination and remyelination. Moreover, MPRAGE is a sensitive and non invasive method that allows to arrange longitudinal studies and to create a bridge between animal and human studies. These data may help to develop new biomarkers and therapeutic targets to treat demyelinating diseases. References 1. Franklin RJ, Ffrench-Constant C. Nat Rev Neurosci. 2008;9(11):839–55. 2. Ludwin SK. Ann Neurol 1994;36(Suppl):S143–5. Review.
|Titolo:||Brain arachidonic acid metabolic pathway is altered during cuprizone-induced demyelination|
|Anno del prodotto:||2009|
|Appare nelle tipologie:||4.2 Abstract in Atti di convegno|